Synergistic induction of nitric oxide synthase type II -: In vitro effect of leptin and interferon-γ in human chondrocytes and ATDC5 chondrogenic cells

Objective. To study, in vitro, the effect of leptin (OB), alone or in combination with interferon-gamma (IFNgamma), on inducible nitric oxide synthase (iNOS) and NO production in human primary chondrocytes and in mouse embryonic chondrogenic ATDC5 cells. Methods. Leptin receptor expression and iNOS messenger RNA expression were evaluated by reverse transcriptase-polymerase chain reaction. Then, iNOS activity was indirectly studied by measuring nitrite accumulation, using the Griess colorimetric reaction, in culture medium of human primary chondrocytes and ATDC5 cells. Results. ATDC5 mouse embryonic cells expressed functional OB receptor. Alone, neither OB nor IFNgamma produced nitrite accumulation in culture medium. However, costimulation with OB and IFNgamma resulted in dose-dependent up-regulation of the expression of iNOS and NO production in human primary chondrocytes and ATDC5 cells. Production of NO was blunted by the iNOS-specific inhibitors L-N-G-nitroarginine methyl ester and aminoguanidine. In addition, the janus-activated kinase 2 (JAK2)-specific inhibitor Tyrphostin AG 490 completely blocked OB + IFNgamma-driven up-regulation of iNOS and NO production. Conclusion. Our data show for the first time a putative proinflammatory role of OB via iNOS induction and NO production. This occurs via activation of JAK2.